Determination of chloramphenicol in milk powder using liquid-liquid cartridge extraction (Chem Elut) and liquid chromatography-tandem mass spectrometry.

BACKGROUND: The European Union prohibits the use of chloramphenicol (CAP) as a veterinary drug in food-producing animals. Nevertheless, CAP have been detected in milk products (liquid milk and milk powder). Therefore, it is necessary to develop sensitive methods for determining CAP residues in milk powder. OBJECTIVE: The aim of this study was to develop and validate a confirmatory method for determination of CAP in milk powder. MATERIAL AND METHODS: Chloramphenicol was determined in milk powder using LC-ESI-MS/MS in negative mode. After fat removing milk powder sample was extracted/cleaned-up with a Chem Elut extraction cartridge. Separation was achieved on a Phenomenex Luna C-18 column with acetonitrile-water as a mobile phase. The mass spectrometer was operated in multiple reaction monitoring mode (MRM). Four transitions were monitored m/z 321→152, 321→194, 321→257 (CAP) and 326→157 (IS CAP-d5). RESULTS: Linearity, accuracy, precision, decision limit (CCa), detection capability (CCb) and ruggedness were determined for m/z 321→152. The mean relative recoveries (inter standard-corrected) of CAP from whole milk powder spiked at levels 0.1, 0.2, 0.3 and 0.6 mg/kg were in the range 95 - 103%. Relative standard deviation (RSD%) of recoveries at all spiked levels were less than 14%. RSDs within-laboratory reproducibility calculated at fortification of 0.3 mg/kg was less than 16%. CCa and CCb were below 0.1 mg/kg. CONCLUSIONS: The developed LC-MS/MS method allows the determination of CAP in milk powder. The method was validated according to the Commission Decision No. 2002/657/EC requirements. This method can be applied to determination CAP in whole and skim milk powder.

Development and validation of a new method for determining nitrofuran metabolites in bovine urine using liquid chromatography - tandem mass spectrometry.

BACKGROUND: The use of nitrofurans as veterinary drugs in food-producing animals is banned throughout the European Union. Nevertheless, nitrofuran metabolites have been detected not only in animal products, but also in bovine urine. At present there are no methods yet published for the simultaneous detection of nitrofuran metabolites in bovine urine. OBJECTIVES: To develop and validate a method for determination of four key nitrofuran metabolites in bovine urine. MATERIAL AND METHODS: The four nitrofuran metabolites (nitrofurantoin, furazolidone, nitrofurazone and furaltadone), were determined in bovine urine using LC-ESI-MS/MS. The procedure required an acid-catalysed release of protein-bound metabolites, followed by their in situ conversion into 2-nitrobenzaldehyde (NBA) derivatives. The sample clean-up was performed using a polymer extraction cartridge before hydrolysis. Nitrofuran metabolites were then determined using electrospray ionization in the positive mode, that had previously been separated on a Phenomenex Luna C-18 column. RESULTS: The method was validated in accordance with the procedure outlined in the Commission Decision No. 2002/657/ EC. Urine samples were spiked with nitrofuran metabolite solutions at levels of 0.5, 1.0, 1.5 and 2.0 microg/kg. Recoveries ranged between 90 - 108% (inter standard-corrected), with a repeatability precision (RSD) of less than 19% for all four analytes. The decision limit (CC) and detection capability (DC) were obtained from a calibration curve and lay respectively within the following ranges: 0.11 - 0.34 microg/kg and 0.13 - 0.43 microg/kg. CONCLUSIONS: The developed and validated LC-ESI-MS/MS method allows four nitrofuran metabolites to be identified and quantitated in bovine urine. This analytical procedure meets the criteria defined in the Commission Decision No. 2002/657/EC.

Increased concentration of metronidazole and its hydroxy metabolite in colon cancer in women.

BACKGROUND: Metronidazole (MTZ) is indicated in the prevention of infections during surgical procedures. However, some data have shown that metronidazole has carcinogenic potential. METHODS: In the present work, we determined concentrations of metronidazole and its hydroxy metabolite (MTZOH) in colorectal cancer patients. MTZ and MTZOH were measured in tumor tissue and surrounding healthy tissue by LC-ESI-MS-MS method. RESULTS: We found different concentration of MTZ and MTZOH in colorectal cancer and healthy tissue. Interestingly, we noted a higher level of the above substances in women vs. men, both in healthy and cancerous gut. CONCLUSION: We suggest that women are more exposed to a potential carcinogenic effect of metronidazole than men.

[Determination of the thyreostats in animal muscle tissue by matrix solid-phase dispersion (MSPD) and liquid chromatography-tandem mass spectrometry]. / Zastosowanie metody ekstrakcji rozpraszania próbki w fazie stalej (MSPD) do oznaczania tyreostatyków w tkance miesniowej zwierzat metoda chromatografii cieczowej-tandem spektometrii mas.

BACKGROUND: The residues ofthyreostats must not be present in the edible animal tissues. The proposed in the EU minimum required performance limit (MPRL) in the animal tissues is 10 microg/kg. This implies the decision limit (CCalpha) and decision capability (CCbeta) of the analytical methods used for the determination of these compounds lower than 10 microg/kg. OBJECTIVE: This study aimed at the development, basing on the literature data and own studies the analytical method allowing for the identification and quantification of five thyreostats: tapazole (TAP), thiouracil (TU), methylotiouracil (MTU), propylothiouracil (PTU) and phenylotiouracil (FTU)) in the bovine muscle tissue, which would meet the criteria set in the Commission Decision No 2002/657/EC. MATERIAL AND METHODS: The developed method used liquid chromatography-tandem mass spectrometry (LC-MS/MS). The sample was extracted and cleaned using the matrix solid-phase dispersion (MSPD) method. The LC was equipped with column Luna C18 Phenomenex. Dimetylotiouracyl was used as internal standard. The samples were fortified at levels: 5, 10 and 20 microg/kg. The method was validated according to the criteria laid down in Commission Decision No. 2002/657/EC. RESULTS: At the levels, mean relative recoveries was in the range 90 - 109% and repeatability (CV %) was less than 10%. Decision limit (CCalpha) and detection capability (CCbeta) calculated for all thyreostats were below the recommended minimum required performance limit (MRPL) - 10 microg/kg. CONCLUSIONS: The developed and validated LC-ESI-MS/MS method allows for the identification and quantification of five thyreostats in the bovine muscle tissue in the quantities below 10 microg/kg. Analytical procedure meets the criteria of Commission Decision No 2002/657/EC.

[Determination of chloramphenicol residues in milk powder using molecular imprinted polymers (MIP) by LC-MS/MS]. / Zastosowanie polimerów z odwzorowaniem czasteczkowym do oznaczania pozostalosci chloramfenikolu w mleku w proszku metoda LC-MS/MS.

The method is presented to analyze chloramphenicol in milk powder. Sample was clean-up by used molecular imprinted polymers (MIP). The determination was performed by LC-ESI-MS/MS. The LC was equipped with column Luna C18 Phenomenex. CAP-d5 was used as internal standards. The method was validation according to the criteria of Decision Commission No 2002/657/EC. Recoveries for the level 0.3 microg/kg was in the range 104-111%. The limit of decision (CCalpha) and detection capability (CCbeta) was respectively 0.06 microg/kg and 0.09 microg/kg.

[Determination of azaperone and carazolol residues in animals kidney using LC-MS/MS method]. / Oznaczanie pozostalosci azaperonu i karazololu w nerkach zwierzat metoda LC-MS/MS.

The method is presented to analyze azaperone and carazolol in pigs kidney. Samples were extracted with acetonitryle. The determination was performed by LC-ESI-MS/MS. The LC was equipped with column Luna C18 Phenomenex. Haloperidol was used as internal standards. The method was validation according to the criteria of Decision Commission No 2002/657/EC. Recoveries for the level 100 microg/kg azaperone and 25 microg/kg carazolol were in the range 91.2-107.0% and 70.8-93.2%. The limit of decision (CCalpha) and detection capability (CCbeta) was respectively 125.9 microg/kg, 160.0 microg/kg for azaperone and 29.3 microg/kg, 33.3 microg/kg. LC-MS/MS technique fulfill the requirements of Decision Commission No 2002/657/EC.

[Determination of malachite green and leukomalachite green residues in fish muscle by high-performance liquid chromatography method]. / Oznaczanie pozostalosci zieleni malachitowej i leukomalachitowej w tkance miesniowej ryb metoda wysokosprawnej chromatografii cieczowej.

Malachite green (MG) and leukomalachite green (LMG) are subjected to monitoring fish muscle, with a minimum required performance limit (MRPL) set 2 microg/kg. Samples were extracted with acetonitryle-buffer mixture and cleaned up on SCX solid phase extraction (SPE) column. LC separation MG and LMG was done on column Luna Phenyl-Hexyl Phenomenex. Samples were fortified with MG and LMG between 2 - 25 microg/kg. The coefficients of variation (CV%) were lower than 14% for MG and 16%for LMG. The mean recoveries were in the range 65- 83% for MG and 70- 73% for LMG. This method fulfils the criteria for identification and determination of MG andLMG residues in the samples of fish muscle.

Determination of nitrofuran metabolites in milk by liquid chromatography-electrospray ionization tandem mass spectrometry.

An LC-ESI-MS-MS method for the analysis of metabolites of four nitrofurans (furazolidone, furaltadone, nitrofurazone and nitrofurantoin) in raw milk has been developed. The samples were achieved by hydrolysis of the protein-bound drug metabolites, derivatization with 2-nitrobenzaldehyd (2-NBA) and clean-up extraction liquid-liquid with ethyl acetate. LC separation was achieved by using a Phenomenex Luna C-18 column. The mass spectrometer operated in multiple reaction monitoring mode (MRM) with positive electro-spray interface (ESI). The method validation was done according to the criteria laid down in Commission Decision No. 2002/657 EC. The validation includes the determination of linearity, repeatability, within-laboratory reproducibility, accuracy, decision limit (CCalpha) and detection capability (CCbeta). The calibration curves were linear, with typical (R(2)) values higher than 0.991. The coefficient of variation (CV, %) was lower than 9.3% and the accuracy (RE, %) ranged from -9.0% to 7.0%. CV within-laboratory reproducibility was lower than 13%. The limits of decision (CCalpha) and detection capability (CCbeta) were 0.12-0.29 microg/kg and 0.15-0.37 microg/kg, thus below the minimum required performance limit (MRPL) set at 1 microg/kg by the UE. This validated method was successfully applied for the determination of nitrofuran metabolites in a large number of milk samples.

[Determination of nitrofuran metabolites residues in animal tissues by LC-MS/MS method]. / Oznaczanie pozostalosci metabolitów nitrofuranów w tkankach zwierzat metoda LC-MS/MS.

A LC-MS-MS method is presented to analyze simultaneously the metabolites of four nitrofuran veterinary drugs in animal muscle tissue e.g., furazolidone, furaltadone, nitofurantoina and nitrofurazone. The sample clean up were performed by a liquid-liquid extraction with ethyl acetate, after a hydrolysis and derivatization with 2-nitrobenzaldehyde. Nitrofurane metabolites were determined by LC-ESI-MS/MS in positive mode. The LC was equipped with column Luna C18 Phenomenex. A binary gradient mobile phase was used as methanol solvent B containing 0.5 mM ammonium acetate and methanol (80:20 v/v). The method was validated according to criteria of Decision Commission No 2002/657/EC. Samples were fortified with metabolites of nitrofuran between 0.5-2.0 microg/kg with AOZ-d4, and AMOZ-d5 as internal standard. The mean recoveries from meat spiked at 1.0 microg/kg were 84.5-109.7%. Limit of decision (CCalpha) was between 0.25-0.57 and capability of detection (CCbeta) 0.32-0.77 microg/kg.

[Determination of chloropromazine residues in animals kidney and urine using LC-MS/MS method]. / Oznaczania pozostalosci chloropromazyny w nerkach i moczu zwierzat metoda LC-MS/MS.

Chloropromazina (CP) is subjected to monitoring food animals products, with a minimum required performance limit (MPRL) set 5.0 microg/kg. Homogenized kidney and urine were extracted with acetonitrile. CP- d3 was used as internal standard. LC separation was done on Luna C18 150 x 2 mm, 5 microm column in mobile phase acetonitrile-acetic acid. CP was determination by LC-ESI-MS/MS negative mode. The method was validation according to the criteria of Decision Commission No 2002/657/EC. Recoveries for the level 5.0 ng/g were in the range 84-102%. The limit of decision (CCalpha) and detection capability (CCbeta) CP in kidney were 1.19; 2.87 ng/g and urine 1.08; 2.61 ng/g.

[Determination of chloramphenicol residues in animal tissues by LC-MS/MS method]. / Oznaczanie pozostalosci chloramfenikolu w tkankach zwierzat metoda LC-MS/MS.

A liquid chromatographic method with mass spectrometric determination and identification (LC-MS/MS) for the determination of chloramphenicol (CAP) in tissues of animal origin was presented. Homogenized samples were extracted with ethyl acetate and evaporated to dryness followed by a clean-up using the liquid-liquid distribution between acetonitryle and hexane. Chloramphenicol was determined by LC-MS-ESI-MS in negative mode. There was used Phenomenex column with the mixture of acetonitrile-water as a mobile phase. The method was validated according to the criteria of Decision Commission No 2002/657/EC. Samples were fortified at CAP levels between 0.1 and 0.45 ng/g with 5d-CAP as internal standard. Recoveries for the level 0.3 ng/g were in the range 76.3-100.3%. Limit of decision (CCalpha) was between 0.137-0.205 ng/g. Limit of quantification (LOQ)--0.1 ng/g.